1. Field of the Invention
The present invention is directed to a prophylactic vaccine effective against Neisseria gonorrhoeae (N.g.) and consequent infection therefrom. Specifically, the instant invention is directed to a fragment of pili protein obtained from gonococci, a method of isolating and purifying same, and a vaccine prepared therefrom.
2. Description of the Prior Art
Gonorrhea is an extremely common infection in humans, and in the United States alone, approximately four million persons annually appear in clinics and physicians offices with this sexually transmitted disease. Despite effective antibiotics, the annual number of cases of gonorrhea in the U.S. has remained essentially unchanged for the past five years. Some gonococci have become resistant to treatment with penicillin, which formerly was one of the most frequently employed medicaments, and there is a growing need for a vaccine to contain this contagion.
Heretofore, no successful vaccine has been available having efficacy against the numerous strains of N.g. One of the principal problems which has impeded the production of a vaccine is the antigenic heterogeneity of the N.g. microorganisms. One of the important N.g. fragments that demonstrates the antigenic heterogeneity are the pili, filamentous structures or hair-like organelles that extend from the surface of virulent N.g. organisms to facilitate their attachment to human cells. Pili were first discovered on N.g. in 1971 (Jephcott, A. E., et al.: Acta Pathol. Microbiol. Scand. Sect. B., 79:437-439, June 1971. "Brief Report: Neisseria gonorrhoeae. III. Demonstration of Presumed Appendages to Cells from Different Colony Types."; Swanson, J. S., et al.: J. Exp. med., 134:886-906, October, 1971. "Studies on Gonococcus Infection. I. Pili and Zones of Adhesion: Their Relation to Gonococcal Growth Patterns.") and first purified in 1972 and 1973 (Buchanan, T. M., et al.: J. Clin. Invest. 52:2896-2909, Nov. 1973, "Quantitative Determination of Antibody to Gonococcal Pili."). They were subsequently shown to have antigenic heterogeneity (Buchanan, T. M.: J. Exp. Med. 141:1470-1475, June 1975, "Antigenic Heterogeneity of Gonococcal Pili."; Buchanan, T. M. and Pearce, W. A.: Infect. Immun. 13:1483-1489, May 1976, "Pili as a Mediator of the Attachment of Gonococci to Human Erythrocytes."; Brinton, C. C., Jr., et al.: Immunobiol. Neisseria gonorrhoeae Proc. Conf. (1978):155-178, Jan. 18-20, 1978, "Uses of Pili in Gonorrhea Control: Role of Bacterial Pili in Disease Purification and Properties of Gonococcal Pili, and Progress in the Development of a Gonococcal Pilus Vaccine for Gonorrhea." Amer. Soc. Microbiol. Wash. D.C., Sept. 29, 1978; Buchanan, T. M., et al.: Immunobiol. Neisseria gonorrhoeae Proc. Conf. (1978): 145-154, Jan. 18-20, 1978, "Pili and Principal Outer Membrane Protein of Neisseria gonorrhoeae: Immunochemical, Structural, and Pathogenic Aspects." Amer. Soc. Microbiol., Wash. D.C., Sept. 29, 1978; Buchanan, T. M.: J. Infect Dis. 138:319-325, Sept. 1978, "Antigen-Specific Serotyping of Neisseria gonorrhoeae I. Use of an Enzyme-Linked Immunosorbent Assay to Quantitate Pilus Antigens on Gonococci." The significance of the pili antigenic heterogeneity was implied in chimpanzees (Arko, R. J., et al.: J. Infect. Dis. 130:160-164, August 1974, "Neisseria gonorrhoeae: Effects of Systemic Immunization on Resistance of Chimpanzees to Urethral Infection."), since immunized chimps were protected against a urethral challenge infection if the challenge N.g. organisms contained pili of the same antigenicity as the N.g. organisms in the vaccine, but not if the pili of the vaccine and challenge N.g. were different. Brinton, et al. (1978, supra) used purified whole pili of a single antigenic type to immunize human volunteers, and demonstrated that these volunteers were more protected that non-immunized volunteers when 10.sup.2 N.g. of the same pili type were introduced into their urethra. However, it has been shown that more than 35 different antigenic types of pili exist on gonococci (Brinton, et al, 1978, supra). Consequently, a vaccine containing only a single pilus antigenic type would be expected to protect against less than 3% of infections with N.g., since antibodies prepared to a single pilus antigenic type only maximally protect against attachment mediated by that same antigenic type of pilus.
The attachment capabilities of isolated gonococcal pili were demonstrated in 1976 (Buchanan & Pearce, 1976, supra) and further characterized in 1978 (Pearce, W. A. and Buchanan, T. M.: J. Clin. Invest. 61:931-943, April 1978, "Attachment Role of Gonococcal Pili: Optimum Conditions and Quantitation of Adherence of Isolated Pili to Human Cells in Vitro."), including initial chemical characterization of the receptor on the human cell surface to which pili bind (Buchanan, T. M., et al.: Immunobiol. Neisseria gonorrhoeae Proc. Conf. (1978):242-249, Jan. 18-20, 1978, "Attachment of Neisseria gonorrhoeae Pili to Human Cells, and Investigations of the Chemical Nature of the Receptor for Gonococcal Pili." Amer. Soc. Microbiol., Wash, D.C., Sept. 29, 1978.). Also, the amino acid composition of pili of antigen type 33 (Hermodson, M. A., Chen, K. C. S., and Buchanan, T. M.: Biochemistry 17:442-445, Feb. 1978, "Neisseria Pili Proteins: Amino-Terminal Acid Sequences and Identification of an Unusual Amino Acid.") and of antigen type CDC-B (Brinton, et al. 1978, supra), and the amino acid sequence of the first 29 residues of pili antigenic types F62, B, 33, and 7122 were published in 1978 (Hermodson, Chen & Buchanan, 1978, supra).
Despite this significant progress, a successful vaccine to prevent gonorrhea, based upon intact whole pili of N.g. seemed unlikely. This was because it was demonstrated that intact whole pili of a single antigenic type, when used as a vaccine, produce antibody primarily to that single antigenic type, rather than to shared pili antigens (Buchanan, 1975, supra; Buchanan & Pearce, 1976, supra; Brinton, 1978, supra, Buchanan, p242-244, 1978, supra). These antibodies specific to a single pilus antigenic type were maximally effected only against N.g. containing the same pili antigens. Since there are more than 35 different antigenic types of pili, it is impractical to prepare a vaccine of intact whole pili containing enough antigenic types to provide protection against most N.g. that cause gonorrhea.